Surface plasmon resonance imaging for real-time, label-free analysis of protein interactions with carbohydrate microarrays

Plant lectin recognition of glycans was evaluated by SPR imaging using a model array of N-biotinylated aminoethyl glycosides of beta-D-glucose ( negative control), alpha-D-mannose ( conA-responsive), beta-D-galactose ( RCA(120)-responsive) and N-acetyl-beta-D-glucosamine ( WGA-responsive) printed onto neutravidin-coated gold chips. Selective recognition of the cognate ligand was observed when RCA(120) was passed over the array surface. Limited or no binding was observed for the non-cognate ligands. SPR imaging of an array of 40 sialylated and unsialylated glycans established the binding preference of hSiglec7 for alpha 2-8-linked disialic acid structures over alpha 2-6-sialyl-LacNAcs, which in turn were recognized and bound with greater affinity than alpha 2-3-sialyl-LacNAcs. Affinity binding data could be obtained with as little as 10-20 mu g of lectin per experiment. The SPR imaging technique was also able to establish selective binding to the preferred glycan ligand when analyzing crude culture supernatant containing 10 20 mu g of recombinant hSiglec7-Fc. Our results show that SPR imaging provides results that are in agreement with those obtained from fluorescence based carbohydrate arrays but with the added advantage of label-free analysis.